Regulation of Regenerative Medicines: A Global Perspective
51
Briefly described below are the three most
commonly used viral vectors for gene therapy—
adenoviruses, AAVs, and lentiviruses.
Adenovirus
Adenoviruses are double-stranded DNA viruses,
~80–100 nm in size and non-enveloped, with
an icosahedral capsid structure.13 Information
derived from preclinical and clinical experience
indicates that adenoviral vectors have a benign
safety profile, with key advantages for their use in
gene therapy, such as their ability to infect both
dividing and non-dividing cells as well as lack of
integration into the host genome.14 Compared to
other common viral vector-based systems, 50%
of clinical trials use adenoviruses, while 28% and
22% of clinical trials employ AAVs and lentivi-
ruses, respectively.15
While adenoviral vectors have been regarded
as an efficient delivery system, a key limitation
in their use lies in the transient expression of
transgenes and high immunogenicity. Therefore, in
order to achieve sustained efficacy, repeat adminis-
trations of adenoviral-based gene therapy products
may be required, leading to the induction of a
strong immune response. Efforts to re-engineer
adenoviral vectors to address these limitations and
introduce improvements, such as increased capac-
ity, remain an active area of reasearch.16
Adeno-Associated Virus (AAV)
AAVs are ~26 nm, single-stranded and non-en-
veloped viruses with an icosahedral capsid
structure.17 Seminal research in the 1960s
exploring their biology has allowed for the
continued evolution of their design as vectors in
gene therapy, leading to the first application of
recombinant AAV (rAAV) in the early 1990s
and the first EMA-approved rAAV gene therapy
in 2012. Note: The first FDA-approved rAAV
product, Luxturna, did not receive licensure until
2017.18 Compared to their wildtype counterpart,
rAAVs lack AAV coding sequences. This feature
increases the packaging capacity of rAAVs and
mitigates some immunogenicity and cytotoxicity
in vivo. However, key limitations to the use of
AAVs as gene therapy vectors remain, including
their inability to accommodate 5.0 kb and the
incomplete knowledge of the long-term durabil-
ity of expression.19 A major challenge also lies in
implementing efficient large-scale manufacturing
methods to produce these vectors as well as the
high cost associated with them.
Lentivirus
Like adenoviruses and AAV, a continued under-
standing of the biology of lentiviruses has paved
the way for their increased use in gene therapy.
Lentiviruses are a subtype of retroviruses, which
are ~80–120 nm, single-stranded RNA, spherical,
and enveloped viruses.20 As vectors, lentiviruses
integrate into the genome, allowing long-term
expression of large (up to 9 kb) transgenes and
are capable of transducing both proliferating and
non-proliferating cells. However, a consequence
of long-term gene expression through integration
into the host genome is the risk of malignant
transformation, which mandates close long-term
follow-up of patients who receive lentiviral-based
therapies. While continued modifications to
lentiviral vectors are underway to achieve better
safety profiles, specificity, expression, and trans-
duction efficiency, the process by which lentiviral
vectors are manufactured remains a major hurdle.
Good manufacturing practice (GMP)-compliant
large-scale manufacturing of lentiviral vectors
require complex production processes and robust
purification methods to generate sufficient vector
quantities to meet clinical demand.21
Regulatory Considerations
Preclinical Assessment
In November 2013, CBER provided recommen-
dations to sponsors on the required preclinical
information to support clinical trials for CGT
products under an investigational new drug
(IND) application and biologics license appli-
cation (BLA).22 Adequate pharmacology and
toxicology studies, conducted in vitro and in
in vivo animal models, are needed to ensure
sufficient information is available on the safety
profile of the investigational gene therapy prod-
uct prior to its administration to humans. Given
the inherent complexity of the products them-
selves, the innovative processes by which they
51
Briefly described below are the three most
commonly used viral vectors for gene therapy—
adenoviruses, AAVs, and lentiviruses.
Adenovirus
Adenoviruses are double-stranded DNA viruses,
~80–100 nm in size and non-enveloped, with
an icosahedral capsid structure.13 Information
derived from preclinical and clinical experience
indicates that adenoviral vectors have a benign
safety profile, with key advantages for their use in
gene therapy, such as their ability to infect both
dividing and non-dividing cells as well as lack of
integration into the host genome.14 Compared to
other common viral vector-based systems, 50%
of clinical trials use adenoviruses, while 28% and
22% of clinical trials employ AAVs and lentivi-
ruses, respectively.15
While adenoviral vectors have been regarded
as an efficient delivery system, a key limitation
in their use lies in the transient expression of
transgenes and high immunogenicity. Therefore, in
order to achieve sustained efficacy, repeat adminis-
trations of adenoviral-based gene therapy products
may be required, leading to the induction of a
strong immune response. Efforts to re-engineer
adenoviral vectors to address these limitations and
introduce improvements, such as increased capac-
ity, remain an active area of reasearch.16
Adeno-Associated Virus (AAV)
AAVs are ~26 nm, single-stranded and non-en-
veloped viruses with an icosahedral capsid
structure.17 Seminal research in the 1960s
exploring their biology has allowed for the
continued evolution of their design as vectors in
gene therapy, leading to the first application of
recombinant AAV (rAAV) in the early 1990s
and the first EMA-approved rAAV gene therapy
in 2012. Note: The first FDA-approved rAAV
product, Luxturna, did not receive licensure until
2017.18 Compared to their wildtype counterpart,
rAAVs lack AAV coding sequences. This feature
increases the packaging capacity of rAAVs and
mitigates some immunogenicity and cytotoxicity
in vivo. However, key limitations to the use of
AAVs as gene therapy vectors remain, including
their inability to accommodate 5.0 kb and the
incomplete knowledge of the long-term durabil-
ity of expression.19 A major challenge also lies in
implementing efficient large-scale manufacturing
methods to produce these vectors as well as the
high cost associated with them.
Lentivirus
Like adenoviruses and AAV, a continued under-
standing of the biology of lentiviruses has paved
the way for their increased use in gene therapy.
Lentiviruses are a subtype of retroviruses, which
are ~80–120 nm, single-stranded RNA, spherical,
and enveloped viruses.20 As vectors, lentiviruses
integrate into the genome, allowing long-term
expression of large (up to 9 kb) transgenes and
are capable of transducing both proliferating and
non-proliferating cells. However, a consequence
of long-term gene expression through integration
into the host genome is the risk of malignant
transformation, which mandates close long-term
follow-up of patients who receive lentiviral-based
therapies. While continued modifications to
lentiviral vectors are underway to achieve better
safety profiles, specificity, expression, and trans-
duction efficiency, the process by which lentiviral
vectors are manufactured remains a major hurdle.
Good manufacturing practice (GMP)-compliant
large-scale manufacturing of lentiviral vectors
require complex production processes and robust
purification methods to generate sufficient vector
quantities to meet clinical demand.21
Regulatory Considerations
Preclinical Assessment
In November 2013, CBER provided recommen-
dations to sponsors on the required preclinical
information to support clinical trials for CGT
products under an investigational new drug
(IND) application and biologics license appli-
cation (BLA).22 Adequate pharmacology and
toxicology studies, conducted in vitro and in
in vivo animal models, are needed to ensure
sufficient information is available on the safety
profile of the investigational gene therapy prod-
uct prior to its administration to humans. Given
the inherent complexity of the products them-
selves, the innovative processes by which they